The role of Stl1p in glycerol accumulation in osmotically stressed icewine yeast Saccharomyces cerevisiae K1 V1116

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Abstract

The high sugar concentration in Icewine juice exerts hyperosmotic stress in the
wine yeast causing water loss and cell shrinkage. To counteract the dehydration,
yeast synthesize and accumulate glycerol as an internal osmolyte. In a laboratory
strain of S. cerevisiae, STLl encodes for Stllp, an H+ /glycerol symporter that is
glucose inactivated, but induced upon hyperosmotic stress.
STLl, was found to be a highly upregulated gene in Icewine fermenting cells
and its expression was 25-fold greater than in yeast cells fermenting diluted Icewine
juice, making it one of the most differentially expressed genes between the two
fermentation conditions. In addition, Icewine fermenting cells showed a two-fold
higher glycerol production in the wine compared to yeast fermenting diluted
Icewine juice.
We proposed that Stllp is (1) active during Icewine fermentation and is not
glucose inactivated and (2) its activity contributes to the limited cell growth
observed during Icewine fermentation as a result of the dissipation of the plasma
membrane proton gradient.
To measure the contribution ofStl1p in active glycerol transport (energy
dependent) during Icewine fermentation, we first developed an Stllp-dependent
(14C]glycerol uptake assay using a laboratory strain of S. cerevisiae (BY 4742 and
LiSTLl) that was dependent on the plasma membrane proton gradient and therefore
energy-dependent. Wine yeast K1-Vll16 was also shown to have this energy
dependent glycerol uptake induced under salt stress. The expression of STLl and Stllp activity were compared between yeast cells
harvested from Icewine and diluted Icewine fermentations. Northern blot analysis
revealed that STLl was expressed in cells fermenting Icewine juice but not
expressed under the diluted juice conditions. Glycerol uptake by cells fermenting
Icewine juice was not significantly different than cells fermenting diluted Icewine
juice on day 4 and day 7 of Vidal and Riesling fermentations respectively, despite
encountering greater hyperosmotic stress. Furthermore, energy- dependent
glycerol uptake was not detected under either fermentation conditions.
Because our findings show that active glycerol uptake was not detected in
yeast cells harvested from Icewine fermentation, it is likely that Stllp was glucose
inactivated despite the hyperosmotic stress induced by the Icewine juice and therefore did not play a role in active glycerol uptake during Icewine fermentation.